RPGRIP1L is required for stabilizing epidermal keratinocyte adhesion through regulating desmoglein endocytosis.

Yeon Ja Choi,Jiang Chen,Chuan Qin,Joshua Lewis,Yusuf A Hannun,Christine Laclef,Elizabeth R Snedecor,Andrew P Kowalczyk,Xuming Mao,Sylvie Schneider-Maunoury,Ken-Ichi Takemaru,Richard A Clark,Li Li,Abraham Andreu-Cervera,Peter J Koch,Kenneth R Shroyer,Aimee S Payne,Ning Yang

doi:10.1371/journal.pgen.1007914

Abstract

Cilia-related proteins are believed to be involved in a broad range of cellular processes. Retinitis pigmentosa GTPase regulator interacting protein 1-like (RPGRIP1L) is a ciliary protein required for ciliogenesis in many cell types, including epidermal keratinocytes. Here we report that RPGRIP1L is also involved in the maintenance of desmosomal junctions between keratinocytes. Genetically disrupting the Rpgrip1l gene in mice caused intraepidermal blistering, primarily between basal and suprabasal keratinocytes. This blistering phenotype was associated with aberrant expression patterns of desmosomal proteins, impaired desmosome ultrastructure, and compromised cell-cell adhesion in vivo and in vitro. We found that disrupting the RPGRIP1L gene in HaCaT cells, which do not form primary cilia, resulted in mislocalization of desmosomal proteins to the cytoplasm, suggesting a cilia-independent function of RPGRIP1L. Mechanistically, we found that RPGRIP1L regulates the endocytosis of desmogleins such that RPGRIP1L-knockdown not only induced spontaneous desmoglein endocytosis, as determined by AK23 labeling and biotinylation assays, but also exacerbated EGTA- or pemphigus vulgaris IgG-induced desmoglein endocytosis. Accordingly, inhibiting endocytosis with dynasore or sucrose rescued these desmosomal phenotypes. Biotinylation assays on cell surface proteins not only reinforced the role of RPGRIP1L in desmoglein endocytosis, but also suggested that RPGRIP1L may be more broadly involved in endocytosis. Thus, data obtained from this study advanced our understanding of the biological functions of RPGRIP1L by identifying its role in the cellular endocytic pathway.

Highlights

Retinitis pigmentosa GTPase regulator interacting protein 1-like (RPGRIP1L, known as NPHP8, MKS5, KIAA1005, or Ftm in mouse) is a transition zone protein that has important roles in regulating cilia formation and function [1,2,3,4,5]
We discovered that RPGRIP1L, a protein known to regulate cilia formation and function, is essential for stabilizing desmosomes of skin keratinocytes
Suppressing the Rpgrip1l gene in mice or in keratinocytes disrupted the ultrastructure of desmosomes, and compromised cell-cell adhesion in vivo and in vitro

Summary

Introduction

Retinitis pigmentosa GTPase regulator interacting protein 1-like (RPGRIP1L, known as NPHP8, MKS5, KIAA1005, or Ftm in mouse) is a transition zone protein that has important roles in regulating cilia formation and function [1,2,3,4,5]. RPGRIP1L is essential for hair follicle morphogenesis by regulating primary cilia formation and hedgehog signaling [11]. RPGRIP1L is expressed in interfollicular epidermal keratinocytes, many of which are not ciliated [12], suggesting that RPGRIP1L may exert cilia-independent functions in the skin. Desmosomes are anchoring junctions that are essential for functionalities of tissues that are subjected to constant mechanical stress, such as the skin and the heart. The adhesion function of desmosomal junctions is dependent on the intercellular anchorage of desmogleins and desmocollins

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